IHC image of PD1 staining in normal human tonsil formalin fixed paraffin embedded tissue section*, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab52587 at 5 µg/ml for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with hematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

Sample: Human tonsil cell extract.
Dilution: ab52587 antibody was used as 1/200 in 1x10⁶ cells/tube.
Anti-CD4 antibody was used as 1/200 dilution.
Anti-CD8 antibody was used as 1/200 dilution.

Formaldehyde-fixed, paraffin-embedded human follicular lymphoma tissue stained for PD1 with ab52587 at 1/100 dilution in immunohistochemical analysis.

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Representative immunohistochemical staining results for PD1 using ab52587 at a 1/50 dilution in human formalin-fixed, paraffin-embedded tissue specimens.

Panel A: Normal lung tissue, negative control;

Panel B: Tonsillar tissue, positive control;

Panel C: PD1-negative tumor infiltrating lymphocytes;

Panel D: PD1-positive tumor infiltrating lymphocytes in squamous cell carcinomas).

Image from PMID: 26313362

Immunohistochemical analysis of various soft tissue sarcomas staining PD1 using ab52587 at a 1/50 dilution.

Arrows indicate PD1 positive lymphocytes.

Image from PMID: 24349382

Overlay histogram showing MOLT-4 (Human lymphoblastic leukemia cell line) cells stained with ab52587 (red line). Live cells were incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab52587, 1/100 dilution) for 30 min at 4°C. The secondary antibody used was Alexa Fluor^® 488 goat anti-mouse IgG (H&L) (ab150117) at 1/2000 dilution for 30 min at 4°C.

MOLT-4 (Human lymphoblastic leukemia cell line) cells stained for PD1 (colored green) using ab52587 in ICC/IF.

Cells were fixed with 100% methanol (5 min) at room temperature and incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% Tween-20 for 1 hour at room temperature to permeabilize the cells and block non-specific protein-protein interactions. The cells were then incubated with ab52587 at 10 µg/ml and ab6046 (Rabbit polyclonal to beta Tubulin - Loading Control) at 1 µg/ml overnight at +4°C. The secondary antibodies used were Goat Anti-Mouse IgG H&L (Alexa Fluor^® 488) (ab150117) secondary antibody used at 1 µg/ml (colored green) and Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 594) (ab150080) secondary antibody (pseudo-colored red) used at 2 µg/ml for 1 hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43 µM for 1 hour at room temperature.

Immunohistochemical analysis of human large and locally advanced breast cancers staining PD1 using ab52587.

(Panel a) Low level of PD-1⁺ T cell infiltration.

(Panel b) high level of PD-1⁺ T cell infiltration. (Itu: intratumoral Str: stromal).

Image data from PMID: 27777963

Human tonsil tissue stained for PD1 with ab52587 incubated for 30 mins at a 1/100 dilution in immunohistochemical analysis.

Immunohistochemical analysis of frozen human liver tissue labeling PD1 with ab52587 at 1/50 dilution.

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Expression of markers of T cell differentiation and degree of inflammation in the heart of chronically T. cruzi-infected subjects with severe cardiomyopathy.

A–D, left panel: representative photos of CD45RO, CD27, PD1 and CD57 expression, respectively.

Panel C shown.

Image data from PMID: 25144227